Laccases catalyze the oxidation of phenols, while reducing molecular oxygen to water. They found high potential applications in biobleaching, decolorization and detoxification. Most of the laccases studied are extracellular, and few intracellular laccases have been isolated form the fruiting bodies of mushrooms (i.e. Lentinula edodes, Cantharellus cibarius, Hericium erinaceum. To our knowledge no work has been reported on the biochemical properties of intracellular laccases from  A. subrufescens . Therefore the aim of this work was to purify and characterize the biochemical properties of laccase(s) from the fruiting bodies of A. subrufescens, in order to establish possible differences between extracellular and intracellular laccases.Purification scheme of the enzyme included homogenization, extraction in phosphate buffer (pH 4), centrifugation (2000 rpm for 20 min), ultrafiltration (polyethersulfone membrane treated with ultraviolet light, UV) and gel chromatography (Sephadex G-75). The kinetics of the purified enzyme was studied using different substrates, and biochemical properties evaluated in terms of optimal pH and temperature, molecular weight, isoelectric point, affinity to different substrates as well as inhibition and activation properties.Results indicated that modifying the surface properties of the polyethersulfone UF membrane by using UV, allowed decreasing adsorption of the enzyme onto the membrane surface and increasing the recovery and the specific activity during the isolation and purification steps. The enzyme showed higher affinity for N�N-dimetil-N- diamine followed by ortodianisidine and 2�2-azino-bis diammonium salt (ABTS). In addition, two iso-enzymes of laccases were identified having different molecular weights and optimal temperatures in ABTS. Results from this work suggest that some of the biochemical properties of intracellular laccases from the fruiting bodies of A. subrufescens are different to those reported for the extracellular laccases produced by this strain. Experimental data indicate that even though extracellular and intracellular lacasses come from the same mushroom, they might have different physiological roles.

Purification and characterization of laccases from the fruiting body of <i>Agaricus subrufescens</i>

247th National Meeting (2014)

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Purification and characterization of laccases from the fruiting body of Agaricus subrufescens